In U.S. Pat. No. 3,517,980 of Jun. 30, 1970, Petran and Hadravsky described a confocal tandem scanning reflected light microscope wherein light is passed to a target through a rotating disc containing several thousand very small holes. Such light is then reflected from the target in a pattern of pinpoints of light through holes in the scanning disc that are exactly opposite the illumination holes. Such a microscope improves resolving power and permits observation of objects covered by translucent materials. The embodiments produced and available to date have used scanning discs having hole diameters of about 60 microns, which holes are larger than is desirable for best microscope resolution, contrast discrimination, and thinnest optical slicing capability. The major reason for not producing microscopes with smaller disc holes is the inability to align such microscopes precisely and easily, and to maintain such alignment, using component mounting schemes and adjustment techniques currently embodied.
This present invention relates to confocal tandem scanning reflected light microscopes having scanning disc holes of diameter as small as 20 microns, and more particularly to an improved optical element mounting apparatus, and to an improved method of adjusting such apparatus. Confocal tandem scanning microscopes having very small scanning disc holes are particularly important and useful for real time, high-magnification viewing of, among other things, living bulk translucent tissues (as in the living eye) and the thinnest possible optical slices thereof, with best resolution and contrast discrimination, all a direct result of the presence of smaller scanning disc holes and precise optical path alignment methods.
In current confocal tandem scanning microscopes using 60 micron holes, alignment of optical elements in the light path is critical, difficult and time consuming, and may not be precise enough to obtain best microscope performance, even for the 60 micron holes. In devices currently on the market, the various elements to be adjusted are basically stacked (piggybacked) so that certain adjustments along the stack are inter-related with other adjustments of elements in the stack. The alignment process involves placing external red and green light sources at the illumination area and viewing area of the disc respectively, and attempting to move the interacting components so as to achieve exact registration of all the red and green pinpoints of light, when viewed through the objective lens opening (both the illumination and the viewing areas can be seen superimposed due to the presence of a beamsplitter). If adjustments can be made so that all the red holes can be exactly superimposed on the all the green holes, the microscope is correctly aligned. Such a method is very laborious and somewhat imprecise when the holes are 60 microns in diameter, and is prohibitively laborious and very imprecise when the holes are 20 or 30 microns in diameter, and therefore is not practical for use when the smaller holes are employed.
One other type of confocal tandem scanning microscope that is currently available (called a "one-path" or "one-sided" confocal tandem scanning microscope) uses disc holes in the 20 to 40 micron range, and requires no special alignment, since it projects the illumination and receives it back through the same scanning hole. Such a microscope is less complex and generally less expensive than a full "two-path" microscope as in the original invention, but has the disadvantage of requiring that the beamsplitter be placed in apposition to the opposite surface of the disc, so that the illuminating light passes through the beamsplitter before passing through the disk, thus exposing the eyepiece to a substantially larger amount of stray reflections than in the two- path microscope. A one-path embodiment is most effective where the light (signal) reflected from the target is bright relative to the light from undesired sources (noise), including stray internal reflections. Such a microscope is readily usable in observing integrated circuit "chips" and other high-reflectance or high contrast targets; however, it is less effective where the light reflected from the target is well below ambient light, such as when the target is one of many types of soft biological tissue (e.g., living eyes) or other relatively non-reflective or low contrast materials.